Various scientific and patent publications are referred to herein. Each is incorporated by reference in its entirety.
Aptamers are molecules that assume an appropriate shape “to fit” another molecule as in a lock and key mechanism. Aptamers can be used as diagnostic tools and/or therapeutics. They were first described in connection with a selection process called SELEX in 1990 by Tuerk and Gold (1). SELEX and its derivatives are based on starting with a large pool of molecules and enriching the pool through a series of iterations until the best binders are discovered. While this process has aided in the discovery of a number of high affinity binders, it is also cumbersome from the need to perform successive rounds of selection and amplification, ending with sequencing and testing of the sequenced aptamers (2). It is further complicated in that aptamers discovered through SELEX may not possess the desired attributes. For example, in the recent description of Cooperative Probe Assays and Tentacle Probes, low affinity binders may be preferable over high affinity binders in order to achieve greater specificity (3). Also the aptamers may need to be in a form such that they can undergo conformational changes producing an increase in signal. Thus there is a need for aptamer selection methods which are faster and easier than conventional SELEX and that allow selection of aptamers based on other properties than high affinity.
Aptamer selection on a chip has been suggested (4). This approach has particular appeal due to the fact that a library of aptamers can be screened for selection and counterselection in a matter of hours, yielding not only information about which sequences function as aptamers, but also information on affinities and/or thermodynamic properties. Unfortunately, chip screening is limited due to library size. Until now, incomplete screenings have been performed requiring multiple steps (5) or using aptamers that are unusually small (hexamers) (4). It is conceivable that mathematical models could screen the library if appropriate algorithms were available, reducing the library to a size that could be placed on a chip. But to date, no algorithms exist that can sufficiently enrich the pool. And given that a 100 mer aptamer contains over 1e60 possible structures and even a library of 20mers contains over 1e12 possibilities, present day computers could not perform all of the computations even if such an algorithm were to exist. The present invention has significant utility for chip based aptamer selection and enables the use of an enriched pool of nucleic acid sequences to define protein binding using both monovalent and multivalent constructs.